LKB1 inhibits HPV-associated cancer progression by targeting cellular metabolism.

Liver kinase B1 (LKB1) is mutationally inactivated in Peutz-Jeghers syndrome and in a variety of cancers including human papillomavirus (HPV)-caused cervical cancer. However, the significance of LKB1 mutations in cervical cancer initiation and progress has not been examined. Herein, we demonstrated that, in mouse embryonic fibroblasts, loss of LKB1 and transduction of HPV16 E6/E7 had an additive effect on constraining cell senescence while promoting cell proliferation and increasing glucose consumption, lactate production and ATP generation. Knockdown of LKB1 increased and ectopic expression of LKB1 decreased glycolysis, anchorage-independent cell growth, and cell migration and invasion in HPV-transformed cells. In the tumorigenesis and lung metastasis model in syngeneic mice, depletion of LKB1 markedly increased tumor metastatic colonies in lungs without affecting subcutaneous tumor growth. We showed that HPV16 E6/E7 enhanced the expression of hexokinase-ll (HK-II) in the glycolytic pathway through elevated c-MYC. Ectopic LKB1 reduced HK-II along with glycolysis. The inverse relationship between HK-II and LKB1 was also observed in normal and HPV-associated cervical lesions. We propose that LKB1 acts as a safeguard against HPV-stimulated aerobic glycolysis and tumor progression. These findings may eventually aid in the development of therapeutic strategy for HPV-associated malignancies by targeting cell metabolism.

LKB1 reduces ROS-mediated cell damage via activation of p38.

Liver kinase B1 (LKB1, also known as serine/threonine kinase 11, STK11) is a tumor suppressor mutated in Peutz-Jeghers syndrome and in a variety of sporadic cancers. Herein, we demonstrate that LKB1 controls the levels of intracellular reactive oxygen species (ROS) and protects the genome from oxidative damage. Cells lacking LKB1 exhibit markedly increased intracellular ROS levels, excessive oxidation of DNA, increased mutation rates and accumulation of DNA damage, which are effectively prevented by ectopic expression of LKB1 and by incubation with antioxidant N-acetylcysteine. The role of LKB1 in suppressing ROS is independent of AMP-activated protein kinase, a canonical substrate of LKB1. Instead, under the elevated ROS, LKB1 binds to and maintains the activity of the cdc42-PAK1 (p21-activated kinase 1) complex, which triggers the activation of p38 and its downstream signaling targets, such as ATF-2, thereby enhancing the activity of superoxide dismutase-2 and catalase, two antioxidant enzymes that protect the cells from ROS accumulation, DNA damage and loss of viability. Our results provide a new paradigm for a non-canonical tumor suppressor function of LKB1 and highlight the importance of targeting ROS signaling as a potential therapeutic strategy for cancer cells lacking LKB1.

Liver kinase B1 regulates the centrosome via PLK1.

Liver kinase B1 (LKB1) is a tumor suppressor mutationally inactivated in Peutz-Jeghers syndrome (PJS) and various sporadic cancers. Although LKB1 encodes a kinase that possesses multiple functions, no individual hypothesis posed to date has convincingly explained how loss of LKB1 contributes to carcinogenesis. In this report we demonstrated that LKB1 maintains genomic stability through the regulation of centrosome duplication. We found that LKB1 colocalized with centrosomal proteins and was situated in the mitotic spindle pole. LKB1 deficiency-induced centrosome amplification was independent of AMP-activated protein kinase (AMPK), a well-defined substrate of LKB1. Cells lacking LKB1 exhibited an increase in phosphorylated and total Polo-like kinase 1 (PLK-1), NIMA-related kinase 2 (NEK2), and ninein-like protein (NLP). Overexpression of active PLK1 (T210D) reversed the inhibition of LKB1 on centrosome amplification. In contrast, depletion of PLK1 with siRNA or suppression of PLK1 kinase activity with BTO-1 (5-Cyano-7-nitro-2-benzothiazolecarboxamide-3-oxide) abrogated LKB1 deficiency-induced centrosome amplification. We further characterized that LKB1 phosphorylated and activated AMPK-related kinase 5 (NUAK1 or ARK5) that in turn increased the phosphorylation of MYPT1, enhanced the binding between MYPT1-PP1 and PLK1, and conferred an effective dephosphorylation of PLK1. More importantly, we noted that LKB1-deficient cells exhibited multiple nuclear abnormalities, such as mitotic delay, binuclear, polylobed, grape, large, and micronuclear. Immediate depletion of LKB1 resulted in the accumulation of multiploidy cells. Expression of LKB1 is reversely correlated with the levels of PLK1 in human cancer tissues. Thus, we have uncovered a novel function of LKB1 in the maintenance of genomic stability through the regulation of centrosome mediated by PLK1.

Piperlongumine induces autophagy by targeting p38 signaling.

Piperlongumine (PL), a natural product isolated from the plant species Piper longum L., can selectively induce apoptotic cell death in cancer cells by targeting the stress response to reactive oxygen species (ROS). Here we show that PL induces cell death in the presence of benzyloxycarbonylvalyl-alanyl-aspartic acid (O-methyl)-fluoro-methylketone (zVAD-fmk), a pan-apoptotic inhibitor, and in the presence of necrostatin-1, a necrotic inhibitor. Instead PL-induced cell death can be suppressed by 3-methyladenine, an autophagy inhibitor, and substantially attenuated in cells lacking the autophagy-related 5 (Atg5) gene. We further show that PL enhances autophagy activity without blocking autophagy flux. Application of N-acetyl-cysteine, an antioxidant, markedly reduces PL-induced autophagy and cell death, suggesting an essential role for intracellular ROS in PL-induced autophagy. Furthermore, PL stimulates the activation of p38 protein kinase through ROS-induced stress response and p38 signaling is necessary for the action of PL as SB203580, a p38 inhibitor, or dominant-negative p38 can effectively reduce PL-mediated autophagy. Thus, we have characterized a new mechanism for PL-induced cell death through the ROS-p38 pathway. Our findings support the therapeutic potential of PL by triggering autophagic cell death.

Crystal structure of human macrophage elastase (MMP-12) in complex with a hydroxamic acid inhibitor.

Human macrophage elastase (MMP-12) is a member of the family of matrix metalloproteinases (MMPs) that plays, like other members of the family, an important role in inflammatory processes contributing to tissue remodelling and destruction. In particular, a prominent role of MMP-12 in the destruction of elastin in the lung alveolar wall and the pathogenesis of emphysema has been suggested. It is therefore an attractive therapeutic target. We describe here the crystal structure of the catalytic domain of MMP-12 in complex with a hydroxamic acid inhibitor, CGS27023A. MMP-12 adopts the typical MMP fold and binds a structural zinc ion and three calcium ions in addition to the catalytic zinc ion. The enzyme structure shows an ordered N terminus close to the active site that is identical in conformation with the superactivated form of MMP-8. The S1'-specificity pocket is large and extends into a channel through the protein, which puts MMP-12 into the class of MMPs 3, 8 and 13 with large and open specificity pockets. The two crystallographically independent molecules adopt different conformations of the S1'-loop and its neighbouring loop due to differing crystal packing environments, suggesting that flexibility or the possibility of structural adjustments of these loop segments are intrinsic features of the MMP-12 structure and probably a common feature for all MMPs. The inhibitor binds in a bidentate fashion to the catalytic zinc ion. Its polar groups form hydrogen bonds in a substrate-like manner with beta-strand sIV of the enzyme, while the hydrophobic substituents are either positioned on the protein surface and are solvent-exposed or fill the upper part of the specificity pocket. The present structure enables us to aid the design of potent and selective inhibitors for MMP-12.

[Transbrachial arterial catheter angiography. Technique, indications, complications]. / Die transbrachiale arterielle Katheter-Angiographie. Technik, Indikationen, Komplikationen.

In a large group of patients, most of whom were attending a department of vascular surgery, who underwent arterial digital subtraction angiography (DSA), a transfemoral approach was not possible in 9%. Therefore in 142 cases transbrachial arterial angiography was electively performed with 4-F or 5-F catheters. This method proved to be a safe and reliable alternative, though the examination technique was more demanding and the complication rate somewhat higher. It offers clear advantages over a transaxillary or translumbar approach. Special attention is paid to the role of transbrachial arterial DSA in evaluating the descending aorta and the peripheral outflow in the legs and to its usefulness in certain angioplasty manoeuvres.

[Comparison of 3 latest-generation glucose micro-reflectometers]. / Vergleich von drei Glucose-Kleinreflektometern der neuesten Generation.

Three glucose reflectance meters (Reflolux II = Accu-Chek II, Glucometer II and Hypocount GA) were tested for precision and accuracy when used by medical personnel of a diabetic outpatient department and for self-monitoring at home. In addition, the visual readability of the appropriate reagent strip was checked. All three systems were sufficiently valid for reflectometric reading, while visual evaluation showed a higher deviation. The precision of Accu-Chek II under optimal conditions in the outpatient department was comparable to the precision of laboratory examinations (CV 3.3%). When used by patients themselves, Accu-Chek II and Glucometer II were sufficiently precise (CV 4.8% [corrected] and 5.3% respectively). These devices are recommended for blood glucose self-monitoring.

[Breast biopsies which failed to show cancer (author's transl)]. / Die nicht zum Karzinomnachweis führende Probeexstirpation aus der Mamma (Verlaufsbeobachtungen und Konsequenzen).

A review is presented on 520 patients who had biopsies of the breast which did not show cancer between 1972-1974. These patients are compared to 304 patients whose biopsies definitely showed cancer of the breast and 520 patients who came to the hospital because of well defined gynaecological disease such as uterine fibroids or uterine prolapse. Comparison of these groups regarding age, past history, indications for the biopsy showed that patients with a biopsy without proof of cancer are difficult to follow for diagnosis. Follow-up on these patients is difficult, however, it is important since up to 4% of these patients with previous negative biopsies later have cancer of the breast. A decrease of biopsies in favor of non-surgical diagnosis of breast lesions can only be recommended under optimal conditions. Under other circumstances the risk of cancer of the breast is high enough to justify a liberal indication for breast biopsies.

[The preventive treatment of general risks prior to gynaecological operations (author's transl)]. / Präventive Therapie allgemeiner Risiken vor gynäkologischen Oerationen.

The operative treatment of high risk patients is of increasing importance in gynaecology. The preoperative preventive treatment of latent general problems is an important tool in the prevention of intraoperative and postoperative complications. The experience of the department for gynaecology in Hanover, Germany, during 1976 is reported. Among 1583 major operations 158 were carried out on patients in the risk class III. Guidelines are described for the preventive treatment of cardiac, bronchopulmonary, hypertensive and diabetic risks. The importance of obesity and advanced age is discussed in our cases.

[Germ cell tumours in gonadal dysgenesis (author's transl)]. / Zur Klinik der Keimzelltumoren bei Gonadendysgenesie

In a 17-year-old patient gonadal dysgenesis with a XY-karyotype was diagnosed by laparoscopy and chromosomal analysis. Two years later, the patient came again to the hospital because of a large tumour which proved to be a dysgerminoma. Except dysgerminomas, other germ cell tumours are also found in gonadal dysgenesis, for example gonadoblastomas, which consist of germinal cells, Sertoli-granulosa-cells and interstitial cells. In most cases of gonadal dysgenesis with a germ cell tumour a Y-chromosome is present. The risk of a gonadal tumour in such cases is estimated to be 25%. In gonadal dysgenesis with a Y-chromosome a prophylactic extirpation of the gonads is necessary, which should be combined with a hysterectomy.